The long-term goal of the research proposed in this grant application is to understand the role of calcium influx in cells of the immune system for immunity to infection and as a cause of immunodeficiency. Our central hypothesis is that calcium influx through so-called CRAC channels is required for the function of T cells and thus immunity to infection. We will study patients with inherited defects in CRAC channel function and use mice with genetic deletion of CRAC channel genes in animal models of infection. We previously showed that patients with inherited defects in CRAC channel function suffer from recurrent, life-threatening infections early in life. CRAC channels are encoded by members of the ORAI and STIM family of genes. We identified the first patients with mutations in ORAI1 and STIM1 genes. They suffer from a unique immunodeficiency syndrome that is characterized by severe infections, autoimmunity, muscular hypotonia and defects in tooth formation and sweat gland function. The immunodeficiency in CRAC channel-deficient patients has been attributed to the impaired function of T cells, white blood cells whose activation is dependent on CRAC channels. Mutations in ORAI1 and STIM1 genes abolish calcium influx in immune cells and impair their function including the production of important immune regulatory proteins. Despite these insights gained from studying the function of CRAC channel-deficient immune cells in cell culture systems, we are lacking a deeper mechanistic understanding of how CRAC channels enable T cells and other immune cells to fight infections in living organisms. In addition to studying patients with inherited mutations in CRAC channel genes, we therefore generated genetically engineered mice that lack expression of STIM and ORAI genes in T cells. Using these mice we recently showed that CRAC channels are important for the ability of T cells to mediate autoimmune diseases and to provide immunity to infection. These mice will therefore be ideal tools to study the mechanisms by which CRAC channels provide host defense to infection with viruses as well as fungal and mycobacterial pathogens. Our lab is in a unique position to translate findings made in animal models into patients by investigating if defects in the calcium-dependent immunoregulatory mechanisms found in mice contribute to the immunodeficiency in patients with mutations in CRAC channels. The specific aims of this proposal are as follows: (1). We will analyze the genetic defects in immunodeficient patients with suspected CRAC channel dysfunction and investigate how mutations interfere with CRAC channel function at a molecular level. (2) We will determine the mechanisms by which CRAC channels in T cells control immunity against acute and chronic viral infections as well as intracellular bacteria. (3) We will investigate the role of CRAC channels in host defense against fungal infections and in regulating the function of T cells that provide antifungal immunity. Furthermore, we will determine how CRAC channels in T cells control immune responses to chronic mycobacterial infections and provide protection against tuberculosis. PUBLIC HEALTH RELEVANCE: The goal of this proposal is to understand the molecular and immunological mechanisms by which calcium influx through CRAC channels in immune cells controls immune responses to infections with viruses, mycobacteria and fungal pathogens. The results from our studies will provide important insights into how calcium influx in immune cells enables them to protect the human host against acute and chronic infection. Since calcium influx is also required for the ability of immune cells to cause autoimmunity, inflammation and allergic responses, CRAC channels have become an attractive drug target to treat these diseases; a better understanding of the role of calcium influx for immunity to infection, however, is required to properly assess the benefits and risks associated with therapeutic CRAC channel inhibition.